Alcoholic liver disease (ALD)is an importanthealthproblem inthe United States. The major difficulty in devising specific therapies for ALD has been our limited understanding of the mechanisms of this liver injury and its systemic complications. This proposal will evaluate the interactionsof cytokines and nutritional/metabolic abnormalities in the development of ALDusing novel animal and in vitro models. Cytokines are low molecular weight mediators of cellular communication that are produced and released by numerous cell types such as monocytes, macrophages, andof particular relevance to liver disease, Kupffer cells and hepatocytes. Pro-inflammatorycytokines such as tumor necrosisfactor-alpha (TNF) and interleukin-18 (IL-18), and chemokines such as interleukin-8 (IL-8) have been postulated to play a role in mediating manyof the systemic manifestations of ALDand the hepatotoxicity in ALD. Itis our working hypothesis that cytokines such as TNF in conjunction with certain metabolic abnormalities observed in ALD,such as altered proteasome function and cAMP metabolism, play an etiologic role inthe development and progression of liver injury in ALD. We postulate that chronic alcohol abusecauses increased gut permeability and endotoxemia(increased LPS), depletionof critical antioxidants (e.g., GSH and Vitamin E),generation of reactive oxygen intermediates, altered methionine metabolism, decreased cAMP, activation of Kupffer cell NFtcB with increased TNFproduction, mitochondrial dysfunction with mitochondrial GSH depletion, proteasome dysfunction, alterations in liver membrane lipid raft composition, increased susceptibility to hepatic TNF cytotoxicity, and liver injury.With decreased proteasome function and hepatocyte injury/death, there is increased IL-8 productionwith neutrophil infiltration andincreasedIL-18 production that sustainsthis inflammatorycytokine response. The focus of our research group has been the interactions of cytokines, oxidative stress, and factors that sensitize to TNF mediated hepatocyte death,such as proteasome dysfunction, in the inductionof ALD. The long-termgoals of our laboratoryare to further define the mechanism(s) involved in the development/evolution of ALDand to develop specific therapies based on this knowledge. The specific objectives of this proposal are: 1. Evaluate the role of cAMP and phosphodiesterase inhibitors on LPS-stimulated TNFproduction in ALD;2: Evaluate monocyte/Kupffer cell Toll-like receptor signaling in ALD;3: Evaluate the effects of hepatocyte lipid rafts on TNF-inducedapoptosis and NFicB activation; and 4: Determinethe effects of proteasome inhibition on "sensitization" to TNF hepatotoxicity and to hepatocyte production of proinflammatory cytokines/chemokines.